Abstract Spinal Muscular Atrophy (SMA) is the leading genetic cause of premature death in children. SMA patients gradually succumb to muscle paralysis as a result of massive motor neuron degeneration. Patients with severe and intermediate forms of SMA experience lifelong physical disability and markedly shorter lifespans, with death as soon as one year (in severe patients). In Dec. 2016, the FDA approved the first and only treatment for SMA, which utilizes intrathecally-administered antisense oligonucleotides (ASOs) upon SMA diagnosis, usually after motor symptoms appear well into the disease progression. ASOs increase SMN (Survival of Motor Neuron) protein expression, which is deleteriously low in SMA patients and is the underlying cause of motor neuron degeneration. ASOs seem to improve motor function and lifespan in patients, but clinical data are still being collected to conclusively determine ASO benefits in humans. Evaluation of ASO efficacy in SMA mouse models support these clinical conjectures, however. Treated mice have improved motor function and longer lifespans, though they remain weaker than healthy littermates and still succumb to eventual motor neuron degeneration and premature death. This is likely because ASO therapy occurs well after the first deficits are established, and central administration of ASO treatment may have reduced access to the peripheral neuromuscular synapses. Without treatment, low levels of SMN produce severe neuromuscular defects that result in up to 50% depression in neurotransmission in SMA model mice. Structural defects include reduced calcium channel expression and clustering (MN cultures & SMA model mice) and fewer neurotransmitter release sites (SMA model mice) in neuromuscular junctions. These deficits contribute to reduced calcium entry into motor nerve terminals (SMA model zebrafish, preliminary data) and reduced calcium transients in growth cones (MN cultures). Because neurotransmission is a calcium-triggered process, these reductions in calcium channels and calcium entry likely explain transmission deficits in SMA model systems. We hypothesize that defective calcium entry underlies reduced neurotransmission in SMA neuromuscular synapses, and that ASO treatment incompletely rescues reduced neurotransmission. This lingering dysfunction ultimately leads to muscular weakness, which precedes motor neuron degeneration. Therefore, an SMN-independent approach that directly targets neuromuscular function by increasing neurotransmission would complement current therapy. We propose to investigate a combination of ASOs (to prolong motor neuron degeneration and overall survival) plus a calcium channel agonist combined with a potassium channel blocker (to increase neurotransmission). We will evaluate these treatments by measuring 1) calcium entry using fluorescent calcium imaging, 2) transmitter release using electrophysiology, and 3) motor function using strength assays.